SUMMARY ? PROJECT 1: SIAH1 IN ER AND MITOCHONDRIAL FUNCTION AND HOMEOSTASIS IN MELANOMA Growing evidence indicates that activation of the adaptive unfolded protein response (UPR) occurs in melanoma cells following drug therapy. The UPR senses oxidative stress, mitochondrial dysfunction and dynamics and resolve protein misfolding in the ER, activities often deregulated in melanoma tumor cells. Notably, the degree of UPR signaling differs among melanoma subtypes, which is consistent with tumor heterogeneity and plasticity and underlies the propensity to adapt to chemotherapy and related stress stimuli. More recently, tumor resistance has been associated with altered expression and activity of the master regulators PGC1? and MITF, factors central for mitochondrial and melanocyte/melanoma biogenesis, respectively. Recent studies, including our preliminary results, demonstrate that deregulation of effectors of the UPR, including ATF4, CHOP and IRE1?, plays a fundamental role in melanoma. Work supported by this P01 established that the ubiquitin ligases Siah1/2 are critical to amplify UPR signaling and identified Siah1 isoform 2 (Siah1is2) as the major isoform induced by the BRAFi (PLX4032) and correlated with PGC1? and MITF expression. Our emerging model is that Siah1is2 is a key sensor that finely tunes the ATF4?IRE1? regulatory axis to control PGC1? and MITF activities, key elements in the propensity of melanoma to adapt to environmental conditions associated with drug resistance and metastatic capacity. Efforts coordinated with Projects 2 and 3 and Cores B and C will further define regulation of PGC1? and MITF expression and activity and evaluate the contribution of ATF4, CHOP, and Siah1is2 to therapeutic responses. To test the hypothesis that UPR- and Siah1is2-controlled signaling engages and fine-tunes the PGC1? and MITF regulatory network to alter metabolic and transcriptional programs, which underlie melanoma resistance and metastasis. We proposed to determine (i) how does Siah1is2 control of the ATF4?PGC1??MITF regulatory axis define melanoma resistance and metastatic phenotypes and (ii) how does ATF4/CHOP regulate melanoma metastasis and drug resistance? Our studies will rely on congenic melanoma tumors, naive or resistant to therapy, congenic Braf/Pten derived melanoma lines CRSPER'd for UPR and Siah1is2 genes, and genetic melanoma models crossed with Atf4, Chop or Siah1 mutant animals. Our studies are expected to lead to unprecedented new insight into the precise role select UPR components play in control of melanoma plasticity, underlying its propensity to resistance and metastatic phenotypes?which represent a key unmet clinical need.